Expression Analysis by Quantitative PCR
Expression analysis often performed in order to monitor changes in gene expression, such as in drug response studies. Quantitative PCR offers a highly sensitive method for determining steady-state mRNA levels present in a sample.
Clients may send RNA or frozen tissues or cell pellets to be extracted. MDS will perform a reverse transcription reaction to generate cDNA, as well as a no reverse transcription control. MDS will use an assay specific to the target to determine the amount present in each sample. Our clients also have a choice of using one of MDS's Optimized Assays, or have MDS develop an assay specific to their target.
The samples can be analyzed using either a relative or absolute standard. A relative standard could be a specific sample that would be used on all plates of the study. An absolute standard could be cDNA made from an in vitro synthesized RNA that contains the target, diluted into an appropriate RNA background. In either case, MDS recommends also running an endogenous control assay to normalize for the amount of starting RNA, such as GAPDH
The sample RNA will be diluted to the appropriate concentration based on information provided by the client. We recommend at least duplicates per sample to be assayed using the target-specific and normalization assays. A DNA positive control will be run on each plate and negative control reactions will include no reverse transcription controls for each sample and a no template control.
- Human P53 gene
- Human PDE7 (phosphodiesterase 7) gene
- Mouse VEGF receptor gene
- Mouse DHFR gene
- Rhesus monkey CFTR gene
- GAPDH (mouse, rabbit, rhesus monkey, hamster, human)
- High copy targets (CHO and human)
Human Cytokines (RNA only)
- IL-1 alpha
- IL-1 beta
- IFN gamma
- TNF alpha